Although some clinical studies have reported increased mitochondrial respiration in patients with fatty liver and early non‑alcoholic steatohepatitis (NASH), there is a lack of in vitro models of non‑alcoholic fatty liver disease (NAFLD) with similar findings. Despite being the most commonly used immortalized cell line for in vitro models of NAFLD, HepG2 cells exposed to free fatty acids (FFAs) exhibit a decreased mitochondrial respiration. On the other hand, the use of HepaRG cells to study mitochondrial respiratory changes following exposure to FFAs has not yet been fully explored. Therefore, the present study aimed to assess cellular energy metabolism, particularly mitochondrial respiration, and lipotoxicity in FFA‑treated HepaRG and HepG2 cells. HepaRG and HepG2 cells were exposed to FFAs, followed by comparative analyses that examained cellular metabolism, mitochondrial respiratory enzyme activities, mitochondrial morphology, lipotoxicity, the mRNA expression of selected genes and triacylglycerol (TAG) accumulation. FFAs stimulated mitochondrial respiration and glycolysis in HepaRG cells, but not in HepG2 cells. Stimulated complex I, II‑driven respiration and β‑oxidation were linked to increased complex I and II activities in FFA‑treated HepaRG cells, but not in FFA‑treated HepG2 cells. Exposure to FFAs disrupted mitochondrial morphology in both HepaRG and HepG2 cells. Lipotoxicity was induced to a greater extent in FFA‑treated HepaRG cells than in FFA‑treated HepG2 cells. TAG accumulation was less prominent in HepaRG cells than in HepG2 cells. On the whole, the present study demonstrates that stimulated mitochondrial respiration is associated with lipotoxicity in FFA‑treated HepaRG cells, but not in FFA‑treated HepG2 cells. These findings suggest that HepaRG cells are more suitable for assessing mitochondrial respiratory adaptations in the developed in vitro model of early‑stage NASH.

• Klíčová slova
• HepG2 cells, HepaRG cells, in vitro models, lipotoxicity, mitochondria, mitochondrial respiration, non‑alcoholic fatty liver disease, steatosis,
• MeSH
• buněčné linie MeSH
• buňky Hep G2 MeSH
• dýchání MeSH
• kyseliny mastné neesterifikované MeSH
• lidé MeSH
• mitochondrie MeSH
• nealkoholová steatóza jater * MeSH
• triglyceridy MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• kyseliny mastné neesterifikované MeSH
• triglyceridy MeSH

Epigallocatechin gallate (EGCG) is a green tea antioxidant with adverse effects on rat liver mitochondria and hepatocytes at high doses. Here, we assessed whether low doses of EGCG would protect these systems from damage induced by tert-butyl hydroperoxide (tBHP). Rat liver mitochondria or permeabilized rat hepatocytes were pretreated with EGCG and then exposed to tBHP. Oxygen consumption, mitochondrial membrane potential (MMP), and mitochondrial retention capacity for calcium were measured. First, 50 μM EGCG or 0.25 mM tBHP alone increased State 4 Complex I-driven respiration, thus demonstrating uncoupling effects; tBHP also inhibited State 3 ADP-stimulated respiration. Then, the coexposure to 0.25 mM tBHP and 50 μM EGCG induced a trend of further decline in the respiratory control ratio beyond that observed upon tBHP exposure alone. EGCG had no effect on MMP and no effect, in concentrations up to 50 μM, on mitochondrial calcium retention capacity. tBHP led to a decline in both MMP and mitochondrial retention capacity for calcium; these effects were not changed by pretreatment with EGCG. In addition, EGCG dose-dependently enhanced hydrogen peroxide formation in a cell- and mitochondria-free medium. Conclusion. Moderate nontoxic doses of EGCG were not able to protect rat liver mitochondria and hepatocytes from tBHP-induced mitochondrial dysfunction.

• MeSH
• hepatocyty cytologie   účinky léků   metabolismus   MeSH
• jaterní mitochondrie účinky léků   metabolismus   MeSH
• katechin analogy a deriváty   farmakologie   MeSH
• krysa rodu Rattus MeSH
• kultivované buňky MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• peroxid vodíku metabolismus   MeSH
• potkani Wistar MeSH
• spotřeba kyslíku účinky léků   MeSH
• terc-butylhydroperoxid toxicita   MeSH
• vápník metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• epigallocatechin gallate MeSH Prohlížeč
• katechin MeSH
• peroxid vodíku MeSH
• terc-butylhydroperoxid MeSH
• vápník MeSH

A compound with promising anticancer properties, 3-bromopyruvate (3-BP) is a synthetic derivative of a pyruvate molecule; however, its toxicity in non-malignant cells has not yet been fully elucidated. Therefore, we elected to study the effects of 3-BP on primary hepatocytes in monolayer cultures, permeabilized hepatocytes and isolated mitochondria. After a 1-h treatment with 100 μM 3-BP cell viability of rat hepatocytes was decreased by 30 % as measured by the WST-1 test (p < 0.001); after 3-h exposure to ≥200 μM 3-BP lactate dehydrogenase leakage was increased (p < 0.001). Reactive oxygen species production was increased in the cell cultures after a 1-h treatment at concentrations ≥100 μmol/l (p < 0.01), and caspase 3 activity was increased after a 20-h incubation with 150 μM and 200 μM 3-BP (p < 0.001). This toxic effect of 3-BP was also proved using primary mouse hepatocytes. In isolated mitochondria, 3-BP induced a dose- and time-dependent decrease of mitochondrial membrane potential during a 10-min incubation both with Complex I substrates glutamate + malate or Complex II substrate succinate, although this decrease was more pronounced with the latter. We also measured the effect of 3-BP on respiration of isolated mitochondria. ADP-activated respiration was inhibited by 20 μM 3-BP within 10 min. Similar effects were also found in permeabilized hepatocytes of both species.

• Klíčová slova
• 3-bromopyruvate, Hepatocyte, Liver, Mitochondria, Toxicity,
• MeSH
• časové faktory MeSH
• hepatocyty cytologie   účinky léků   ultrastruktura   MeSH
• jaterní mitochondrie účinky léků   MeSH
• krysa rodu Rattus MeSH
• kultivované buňky MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• mitochondriální nemoci chemicky indukované   patofyziologie   MeSH
• myši MeSH
• pyruváty farmakologie   toxicita   MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• viabilita buněk účinky léků   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• bromopyruvate MeSH Prohlížeč
• pyruváty MeSH
• reaktivní formy kyslíku MeSH

Epigallocatechin-3-gallate (EGCG) is the main compound of green tea with well-described antioxidant, anti-inflammatory, and tumor-suppressing properties. However, EGCG at high doses was reported to cause liver injury. In this study, we evaluated the effect of EGCG on primary culture of rat hepatocytes and on rat liver mitochondria in permeabilized hepatocytes. The 24-hour incubation with EGCG in concentrations of 10 μmol/L and higher led to signs of cellular injury and to a decrease in hepatocyte functions. The effect of EGCG on the formation of reactive oxygen species (ROS) was biphasic. While low doses of EGCG decreased ROS production, the highest tested dose induced a significant increase in ROS formation. Furthermore, we observed a decline in mitochondrial membrane potential in cells exposed to EGCG when compared to control cells. In permeabilized hepatocytes, EGCG caused damage of the outer mitochondrial membrane and an uncoupling of oxidative phosphorylation. EGCG in concentrations lower than 10 μmol/L was recognized as safe for hepatocytes in vitro.

• MeSH
• čaj chemie   metabolismus   MeSH
• fluorescenční mikroskopie MeSH
• hepatocyty cytologie   účinky léků   metabolismus   MeSH
• jaterní mitochondrie účinky léků   metabolismus   MeSH
• kaspasa 3 metabolismus   MeSH
• katechin analogy a deriváty   toxicita   MeSH
• krysa rodu Rattus MeSH
• kultivované buňky MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• oxidativní fosforylace účinky léků   MeSH
• potkani Wistar MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• TNF-alfa metabolismus   MeSH
• viabilita buněk účinky léků   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• čaj MeSH
• epigallocatechin gallate MeSH Prohlížeč
• kaspasa 3 MeSH
• katechin MeSH
• reaktivní formy kyslíku MeSH
• TNF-alfa MeSH